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Previous issue date: 2009-05-29 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior-CAPES / In spite of the technological progresses, the understanding of the lignin structural
formation is still matter of scientific investigations. This research aimed to utilize
Eucalyptus urophylla S.T. Blake callus for lignin production. This callus were obtained
from stems segments explants grown in culture medium added with a combination of
the cytokine TDZ and the auxins acid indole-3-acetic (IAA), acid ?-naphthaleneacetic
(NAA) and acid 2,4-dichlorophenoxyacetic (2,4-D). This growth regulators were
utilized either alone or mixtured. For cell suspension production it was utilized callus
obtained in medium culture added with 20?M IAA + 3?M TDZ, after 30 days of
growth in vitro. Once the cells suspension was obtained, the lignin production was
induced by four elicitors: jasmonic acid (JA), NAA, sucrose and control (without
elicitor). It was used a completely randomized design with four replications. Each plot
consisted of an Erlenmeyer with 125 ml of cells suspension culture. The Wiesner test
confirmed the lignin presence in all treatments. In 3 of the 4 replicatons it was
performed another evaluation to the production of DHPs (polymers by oxidative
dehydrogenation) utilizing suspension filtrate added with H2O2, H2O2 + peroxidase and
peroxidase. These new treatments were analyzed through polilignols production
utilizing infrared (IR) and nuclear magnetic resonance of the hydrogen (NMR H). The
suspension filtrate analysis of the 4th replication through ultraviolet (UV), IR, NMR 13C
and NMR H evidenced the production of extra cellular lignin. Of this, the largest
content was obtained in presence of sucrose as elicitor, followed by ANA and AJ. The
cells in suspension increased the cellular wall lignin content in all treatments in relation
to the control and the largest values were with the medium containing sucrose. DHPs
were also analyzed utilizing as mattress the MS medium added of the same elicitors
tested in the cellular suspension phase. For this, it was utilized as precursors the
following alcohols: coniferyl or sinapyl, H2O2 and peroxidase; amd the analyses were
done in RMN H and RMN 13C. The results showed DHPs synthesis of both coniferyl
alcohol (DHP1c, DHP2c and DHP4c) and sinapyl alcohol (DHP2s). Nevertheless, It
was not synthesized DHP in the treatment containing sucrose when the precursor was
the coniferyl alcohol. On the other hand, when the sinapyl alcohol was the precursor,
DHPs were only synthesized in the presence of ANA as elicitor. It was concluded that
sucrose is an appropriate elicitor for the lignin production in cells suspension both at
cellular and extra cellular level. However, this result was not observed in relation to
DHP production. NAA auxin had a better functionality in the DHP2c and DHP2s
formation. These results may be considered a progress in the lignification studies with
the use of E. urophylla cell suspension. Once all these questions were answered and
solved, it will be possible to develop E.urophylla suitable plants with better quality
forest products and able to cause smaller environmental impact in the industrial
processes. / Apesar dos avan?os tecnol?gicos, a compreens?o da forma??o estrutural da lignina
ainda ? alvo de in?meras investiga??es cient?ficas. Esta pesquisa foi realizada com calos
de Eucalyptus urophylla S.T. Blake para a produ??o de lignina. Os calos foram obtidos
a partir de explantes de segmentos caulinares desenvolvidos em meios de cultura
acrescidos de uma combina??o da citocinina TDZ e das auxinas ?cidos: 3-indolac?tico
(AIA), ?-naftalenoac?tico (ANA) e diclorofenoxiac?tico (2,4-D), nas formas isoladas e
conjugadas. Para a produ??o de c?lulas em suspens?o foram utilizados os calos
formados no tratamento contendo 20?M de AIA + 3?M de TDZ, ap?s 30 dias de cultivo
in vitro. Obtidas as c?lulas em suspens?o, a produ??o de lignina foi induzida
empregando-se quatro elicitores: ?cido jasm?nico (AJ), ANA, sacarose e testemunha
(sem o emprego de elicitores). O delineamento experimental utilizado foi o inteiramente
casualizado, 4 repeti??es e um Erlenmeyer contendo 125 ml de cultura de c?lulas em
suspens?o. O teste de Wiesner confirmou a presen?a de lignina, em todos os tratamentos
testados. Em 3 das 4 repeti??es foram realizados subtratamentos para a produ??o de
DHPs (pol?meros por desidrogena??o oxidativa) a partir do filtrado da suspens?o com
H2O2, H2O2 + peroxidase e peroxidase. A an?lise desses subtratamentos foi realizada
pela detec??o da produ??o de polilign?is atrav?s de raios infravermelho (IV) e
resson?ncia magn?tica nuclear do hidrog?nio (RMN H). O filtrado da suspens?o da
repeti??o 4 foi analisado por ultravioleta (UV), IV, RMN 13C e RMN H, sendo
constatada a presen?a de lignina extracelular, com o maior teor sendo observado na
presen?a do elicitor sacarose, seguido do ANA e AJ. As c?lulas em suspens?o
apresentaram aumento no teor de lignina na parede celular em todos os tratamentos em
rela??o ? testemunha e os maiores valores foram com o meio contendo sacarose. Foram
analisadas tamb?m as DHPs tendo como colch?o o meio de cultura MS acrescido dos
mesmos elicitores testados na fase de suspens?o celular. Para isto foram utilizados como
precursores os ?lcoois conifer?lico ou sinap?lico, H2O2 e peroxidase, com a lignina
analisada em RMN H e RMN 13C. Os resultados mostraram que houve s?ntese de DHPs
do ?lcool conifer?lico (DHP1c, DHP2c e DHP4c) e do ?lcool sinap?lico (DHP2s).
Porem quando se utilizou como precursor o ?lcool conifer?lico n?o foram sintetizadas
DHPs no tratamento contendo sacarose como elicitor. E, quando foi utilizado como
precursor o ?lcool sinap?lico, somente foram formadas DHPs na presen?a do elicitor
ANA. Concluiu-se, ent?o, que a sacarose apresentou-se como um elicitor adequado para
a produ??o de lignina nas c?lulas em suspens?o, tanto em n?vel celular quanto
extracelular. Entretanto, isto n?o foi observado em rela??o ? produ??o de DHP. A
auxina ANA teve funcionalidade maior na forma??o de DHP2c e DHP2s. Estes
resultados devem ser considerados como um avan?o nos estudos de lignifica??o com a
utiliza??o de c?lulas em suspens?o de E. urophylla. Quando esses questionamentos
forem identificados e solucionados ser? poss?vel o desenvolvimento de novos
indiv?duos desta esp?cie que conduzam ? produ??o de produtos florestais de melhor
qualidade, com menor impacto ambiental nos processos industriais.
| Identifer | oai:union.ndltd.org:IBICT/oai:localhost:jspui/1211 |
| Date | 29 May 2009 |
| Creators | Monteiro, Maria Beatriz de Oliveira |
| Contributors | Abreu, Heber dos Santos, Carvalho, Ana Cristina Portugal Pinto de, Muniz, Graciela In?s Bolzon de, Latorraca, Jo?o Vicente de Figueiredo, Garcia, Rosilei Aparecida, Arruda, Rosani do Carmo de Oliveira |
| Publisher | Universidade Federal Rural do Rio de Janeiro, Programa de P?s-Gradua??o em Ci?ncias Ambientais e Florestais, UFRRJ, Brasil, Instituto de Florestas |
| Source Sets | IBICT Brazilian ETDs |
| Language | Portuguese |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, info:eu-repo/semantics/doctoralThesis |
| Format | application/pdf |
| Source | reponame:Biblioteca Digital de Teses e Dissertações da UFRRJ, instname:Universidade Federal Rural do Rio de Janeiro, instacron:UFRRJ |
| Rights | info:eu-repo/semantics/openAccess |
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